The goal of these studies is to induce long-term specific unresponsiveness to allografts without the need for chronic immunosuppression. A well- studied protocol, treatment of recipients with a short course of antilymphocyte serum (ALS) and donor bone marrow cells (BMC), has promise in this regard, but only a small percentage of grafts are retained long- term. We have recently observed that if the ALS/BMC, skin- or islet cell- grafted mice are exposed to donor strain neonatal tissue, the length of graft survival is considerably increased. Not only do neonatal grafts survive longer that adult grafts, but if adult-grafted animals are co- transplanted with neonatal tissue, the length of adult graft survival is increased. Additionally, it appears that a more profound state of immunological unresponsiveness is induced by exposure to neonatal, rather than adult, alloantigen. We propose to investigate the mechanisms by which neonatal skin allografts and adult allografts co-transplanted with neonatal donor strain tissue enjoy prolonged survival in ALS-BMS treated mice. Cells recovered from animals bearing long-term surviving grafts will be tested in vivo and in vitro for immunological reactivity. Also, the response of cells from naive animals to neonatal and neonatal plus adult alloantigen will be analyzed in vitro. Immunohistochemical analysis of the development of immunoregulatory cells in the transplanted skin will be carried out, and such cells isolated, to determine their contribution to the induction of unresponsiveness. In a pilot experiment in which a short course of therapy with cyclosporin A has been added to the treatment protocol, strikingly improved survival of neonatal skin allografts has been observed (>80% of grafts have survive >180 d). Therefore, this combined treatment will be extended to attempt to prolong survival of adult grafts co-transplanted with neonatal tissue, and to strongly incompatible recipient/donor combinations, including xenogeneic combinations. Cells that preferentially activate suppressor pathways are known to be present in adult tissues and can be recovered by fractionation, or their activity can be "unmasked" by UV-irradiation of the tissue. Therefore, we will attempt to use such adult-donor derived cells to simulate the immunoregulatory effects of neonatal tissue. The understanding which we will gain of the immunoregulatory potential of neonatal alloantigen should lead to the ability to induce specific, long-term unresponsiveness to allografts from adult donors, and ultimately to application to human transplantation.